title: TGF-beta1 reduces Wilms' tumor suppressor gene expression in podocytes.
abstract: BACKGROUND
  Wilms' tumor suppressor gene (WT1) is essential for normal podocyte function, and transforming growth factor (TGF)-beta contributes to focal segmental glomerulosclerosis (FSGS). We aimed to address whether TGF-beta affects WT1 expression in podocytes.
  
  
  METHODS
  A human podocyte cell line treated with TGF-beta1 and kidneys in Alb/TGF-beta1-transgenic mice were analyzed for WT1 expression.
  
  
  RESULTS
  In cultured podocytes, TGF-beta1 reduced WT1 protein expression determined by western blotting beginning at 8 h and decreased WT1 messenger RNA (mRNA) expression measured by quantitative reverse transcription-polymerase chain reaction beginning at 3 h. Knockdown of Smad4 by small hairpin (sh) RNA partially rescued the TGF-beta1-induced reduction of both WT1 protein and mRNA expressions in the cultured podocytes. TGF-beta1 did not alter luciferase activity of the reporter construct for a human WT1 promoter but reduced that for a human WT1 5' enhancer construct, suggesting that TGF-beta1 may regulate WT1 expression by altering the 5' enhancer activity. In the transgenic mice, WT1 protein expression in podocytes was decreased at 1 and 3 weeks of age, while glomeruloclerosis developed after 3 weeks.
  
  
  CONCLUSIONS
  TGF-beta1 reduces WT1 expression in cultured human podocytes and podocytes in mice before overt glomerulosclerosis begins. The effects are at least partially Smad4 dependent. Our findings identify a novel pathway linking TGF-beta1 to podocyte injury and FSGS. The WT1 reduction may be a useful marker for early podocyte injury.
authors.affiliation: Kidney Disease Section, Kidney Disease Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, USA.
authors.forename: Toru
authors.initials: T
authors.lastname: Sakairi
authors.forename: Yoshifusa
authors.initials: Y
authors.lastname: Abe
authors.forename: Jeffrey B
authors.initials: JB
authors.lastname: Kopp
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002478:Cells, Cultured; D005455:Fluorescent Antibody Technique; D015870:Gene Expression; D005923:Glomerulosclerosis, Focal Segmental / Q000209:etiology* / Q000378:metabolism / Q000473:pathology; D006801:Humans; D007124:Immunoenzyme Techniques; D007678:Kidney Glomerulus / Q000378:metabolism* / Q000473:pathology*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008808:Mice, Inbred CBA; D008822:Mice, Transgenic; D050199:Podocytes / Q000378:metabolism* / Q000473:pathology; D011401:Promoter Regions, Genetic / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D051901:Smad4 Protein / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D053773:Transforming Growth Factor beta1 / Q000502:physiology*; D025721:WT1 Proteins / Q000235:genetics* / Q000378:metabolism
chemical_list: D012333:RNA, Messenger; D034741:RNA, Small Interfering; C497796:SMAD4 protein, human; D051901:Smad4 Protein; D053773:Transforming Growth Factor beta1; D025721:WT1 Proteins